Affiliations: 1: Department of Microbiology and Immunology, Stritch School of Medicine, Loyola University Chicago, Maywood, IL 60153;
2: Center for Genomics and Systems Biology, Department of Biology, New York University, New York, NY 10003;
3: New York University, New York, NY;
4: Stritch School of Medicine, Loyola University Chicago, Maywood, IL

Spores of Clostridiales and Bacillales are encased in a complex series of concentric shells that provide protection, facilitate germination, and mediate interactions with the environment. Analysis of diverse spore-forming species by thin-section transmission electron microscopy reveals that the number and morphology of these encasing shells vary greatly. In some species, they appear to be composed of a small number of discrete layers. In other species, they can comprise multiple, morphologically complex layers. In addition, spore surfaces can possess elaborate appendages. For all their variability, there is a consistent architecture to the layers encasing the spore. A hallmark of all Clostridiales and Bacillales spores is the cortex, a layer made of peptidoglycan. In close association with the cortex, all species examined possess, at a minimum, a series of proteinaceous layers, called the coat. In some species, including Bacillus subtilis, only the coat is present. In other species, including Bacillus anthracis, an additional layer, called the exosporium, surrounds the coat. Our goals here are to review the present understanding of the structure, composition, assembly, and functions of the coat, primarily in the model organism B. subtilis, but also in the small but growing number of other spore-forming species where new data are showing that there is much to be learned beyond the relatively well-developed basis of knowledge in B. subtilis. To help summarize this large field and define future directions for research, we will focus on key findings in recent years.

44.Sandman K, Kroos L, Cutting S, Youngman P, Losick R. 1988. Identification of the promoter for a spore coat protein gene in Bacillus subtilis and studies on the regulation of its induction at a late stage of sporulation. J Mol Biol200:461–473. [PubMed][CrossRef]

165.Henriques AO, Beall BW, Moran CP Jr. 1997. CotM of Bacillus subtilis, a member of the alpha-crystallin family of stress proteins, is induced during development and participates in spore outer coat formation. J Bacteriol179:1887–1897. [PubMed]

Spores of Clostridiales and Bacillales are encased in a complex series of concentric shells that provide protection, facilitate germination, and mediate interactions with the environment. Analysis of diverse spore-forming species by thin-section transmission electron microscopy reveals that the number and morphology of these encasing shells vary greatly. In some species, they appear to be composed of a small number of discrete layers. In other species, they can comprise multiple, morphologically complex layers. In addition, spore surfaces can possess elaborate appendages. For all their variability, there is a consistent architecture to the layers encasing the spore. A hallmark of all Clostridiales and Bacillales spores is the cortex, a layer made of peptidoglycan. In close association with the cortex, all species examined possess, at a minimum, a series of proteinaceous layers, called the coat. In some species, including Bacillus subtilis, only the coat is present. In other species, including Bacillus anthracis, an additional layer, called the exosporium, surrounds the coat. Our goals here are to review the present understanding of the structure, composition, assembly, and functions of the coat, primarily in the model organism B. subtilis, but also in the small but growing number of other spore-forming species where new data are showing that there is much to be learned beyond the relatively well-developed basis of knowledge in B. subtilis. To help summarize this large field and define future directions for research, we will focus on key findings in recent years.

Thin-section TEM analysis of spores from diverse species. Spores were prepared as described in McKenney et al. (13). Images in the top row were fixed using ruthenium red. Other images were conventionally fixed. Images are not to scale; each image was sized to facilitate comparison. Two images of Bacillus amyloliquefaciens are shown (one showing a section along the long axis, the other showing a section along the short axis) to point out the thick caps of coat at the poles. The difference in thickness between the two caps is a consistent feature of this species. Two images of Brevibacillus laterosporus are also shown to emphasize the variation in morphology of the distinctive structure (indicated with a brown bracket) associated with the coat. The mother cell envelope, which is still present in these two spores, is indicated with a green bracket. The image of Clostridium difficile is taken from Semenyuk et al. (142). The crust (Cr), outer coat (OC), and inner coat (IC) are indicated in the image of Bacillus subtilis in the upper left. The coat and, where it is present, the exosporium are indicated with blue and red brackets, respectively. The image of Bacillus megaterium is courtesy of Dr. Joel Bozue at the U.S. Army Medical Research Institute of Infectious Diseases (USAMRIID).

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microbiolspec/4/2/TBS-0023-2016-fig1.gif

FIGURE 1

Thin-section TEM analysis of spores from diverse species. Spores were prepared as described in McKenney et al. (13). Images in the top row were fixed using ruthenium red. Other images were conventionally fixed. Images are not to scale; each image was sized to facilitate comparison. Two images of Bacillus amyloliquefaciens are shown (one showing a section along the long axis, the other showing a section along the short axis) to point out the thick caps of coat at the poles. The difference in thickness between the two caps is a consistent feature of this species. Two images of Brevibacillus laterosporus are also shown to emphasize the variation in morphology of the distinctive structure (indicated with a brown bracket) associated with the coat. The mother cell envelope, which is still present in these two spores, is indicated with a green bracket. The image of Clostridium difficile is taken from Semenyuk et al. (142). The crust (Cr), outer coat (OC), and inner coat (IC) are indicated in the image of Bacillus subtilis in the upper left. The coat and, where it is present, the exosporium are indicated with blue and red brackets, respectively. The image of Bacillus megaterium is courtesy of Dr. Joel Bozue at the U.S. Army Medical Research Institute of Infectious Diseases (USAMRIID).

Model of spore coat assembly during B. subtilis sporulation. In the left column, we list the stages of sporulation as they appear by TEM, phase-contrast microscopy, or fluorescence microscopy in the presence of a membrane stain. The center column contains diagrams of spore coat morphogenesis. Layers of the spore coat are color coded (cyan = basement layer; yellow = inner coat; blue = outer coat; maroon = crust). In the right column, we list the stages of spore coat assembly. DPA, dipicolinic acid. Modified from McKenney et al. (21). See text for details.

microbiolspec/4/2/TBS-0023-2016-fig2_thmb.gif

microbiolspec/4/2/TBS-0023-2016-fig2.gif

FIGURE 2

Model of spore coat assembly during B. subtilis sporulation. In the left column, we list the stages of sporulation as they appear by TEM, phase-contrast microscopy, or fluorescence microscopy in the presence of a membrane stain. The center column contains diagrams of spore coat morphogenesis. Layers of the spore coat are color coded (cyan = basement layer; yellow = inner coat; blue = outer coat; maroon = crust). In the right column, we list the stages of spore coat assembly. DPA, dipicolinic acid. Modified from McKenney et al. (21). See text for details.